Apparatus and methods for sensing fluid components

ABSTRACT

Provided herein is a method for accurate, reproducible analytical solution evaluation eliminating the need for a reference sensor by determining the activity of selected species employing species specific sensors and species combination sensors in conjunction with Nernst-type equations. Also provided are sensor structures for elimination of edge effects to signals thereby yielding accurate, reproducible measurements, and a cartridge structure adapted to incorporate an array of the new sensors for employment of the new method where the cartridge is particularly adapted for miniaturization, maintaining a fixed volume of solution for analysis and providing an anaerobic testing environment. Lastly, a compact instrument embodying miniaturization especially adapted for field use and use of the cartridge is provided herein.

This is a continuation of application Ser. No. 008,554, filed 01/29/87now U.S. Pat. No. 1,762,594.

TECHNICAL FIELD

This invention relates to analytical measurement of solutions and,particularly, to a method for referenceless sensor measurement employingsingle point calibration, a new electrode promoting uniform fluxdistribution, a new fixed volume anaerobic sensor cartridge and a newminiaturized instrument for analytical chemical measurements.

BACKGROUND OF THE INVENTION

The traditional "wet" chemistry techniques in analytical chemistry andits more sophisticated progeny, clinical chemistry, have in recentdecades been replaced by electronic instrumentation. With the advent ofinstrumentaton, accuracy in reproducibility of experimental measurementshas been enhanced. Such accuracy is of particular importance in clinicalchemical techniques and of the greatest importance in biomedicalmeasurements where minute (part per million) measurements are common.Linking such instrumentation and automated processing with themicroprocessors and affordable computer technology, has resulted inanother step in the evolution of analytical and clinical chemicaltechniques.

In the sub-discipline of electro-chemical measurements, great advancesin such instrumentation have been made. Generally, conventionalelectro-chemical measurements require the measurement of two samplesolutions containing two different known concentrations of a substancefor calibration purposes followed by measurement of a solutioncontaining an unknown quantity of the species. Electro-chemical methodsgenerally require use of a reference electrode, a substance specificelectrode and a bridge between the solution in order to achieve a cellfor potentiometric measurements. The electrical signal (commonly inmillivolts) obtained from the cell is proportional to the ionic activityand, therefore, concentration of the substance in the solution. Thesignal/concentration relationship is algebraically expressible by aNernst equation

    V=M.sub.f [C]+I+J                                          (1)

where

V is the voltage (signal)

M_(f) is the slope (a constant for the particular electrode andsubstance)

I is a constant for a particular substance

J is the junction potential of the cell

[C] is the ionic activity (concentration of the substance).

In order to establish the values necessary to solve the equation, it isfirst required to determine the slope, M_(f), for the electrode. Forthis step, measurement of two solutions containing known concentrationsare taken, the values inputted into the above equation and the equationssolved simultaneously to obtain the slope. Next, it is necessary todetermine the constant, I, for the particular substance in solutionrelative to the particular electrode. The junction potential is alsodetermined by conventional methods. The foregoing technique is commonlyreferred to as a double or dual point calibration. Recent developmentsin electrode technology have dispensed with the need for the slopedetermination by providing preset, one-shot electrodes where the slopeis known for a particular substance and electrode structure. Thesedevices are generally limited, however, to one time use due to the slopeshift after a period of exposure to solution. Slope shift isattributable to, among other causes, hydration of a previouslyunhydrated electrode. In view of this arrangement, such one shotelectrodes are confined to use with specific systems and particularelectrode arrangements.

Great improvements have been made in the sensitivity of sensors employedfor electro-chemical analysis. Many relatively new sensor types have nowfound their way into the laboratory. Most notable are variants of theion selective electrode (ISE), the enzyme base selective electrode(EBSE), the anti-body based selective electrode (ABSE), chemical fieldeffect transducer (CHEMFET) and the ion selective field effecttransducer (ISFET). Each of these sensor types may be incorporated intoa number of physical variants including coated wire electrodes, thinfilm electrodes, etc. These are employable, not only for clinicalchemical application, but also for general use in such fields asindustrial chemical, pharmaceutical, biochemical, environmental control,etc. Moreover, these devices now provide the technician with aconsiderable selection of devices and techniques which function toproduce electrical signals proportional to the ionic activity of aparticular substance or substances for which the sensors arespecifically designed and, therefore, increasingly precise measurements.

Referring briefly to optical sensors and analytical methods primarilyrelying on colorometry, they, too, have experienced a correspondingrapid evolution. Significant advances are pronounced in the biochemistryfield, e.g. enzyme and antibody-antigen reactions.

However, the technician is now faced with an increasing array ofproblems associated with the new technologies. For example, due to thesensitivity of the above-mentioned sensors, they may possess a bulkydesign. Notably, electro-chemical sensor systems generally require areference electrode and species sensitive electrode, both of which mustbe carefully calibrated or preconditioned. Also, especially in the caseof reference electrodes, supposedly identical electrodes may differslightly due to manufacturing tolerances which can lead to erraticmeasurements, "drift" problems and junction potential errors.

Measurement variations may occur from use of such electrodes due tosignal drift and varying junction potentials between the referenceelectrode in the media being studied and the associated electrodes.Junction potential contribution to the signal not only results fromelectrode structure, but also varies from instrument to instrument aswell as measurement to measurement. For sensitive measurements, suchvariations are wholly unacceptable. Further problems are augmented byincreasing sensitivity of the electrodes, particularly in biomedicalapplications, where precise measurements are critical. Factors such asthe longevity, stability and contamination of the reference electrode,particularly when employed in hostile environments such as invasivemonitoring during surgical procedures, must be accounted for, and have,thus far, escaped resolution. Finally, in devices requiring therelatively bulky reference electrode, electro-chemical systems have, forthe most part, belied miniaturization.

During electro-chemical measurements of complex solutions(multicomponent) another problem arises, namely, separation of thesignal from reference electrode junction potential. For measurement ofcomplex solutions containing many potentially interactive electroactivespecies, in contrast to elementary assessment of a single speciessolution, the coefficient of activity (contribution by individualcomponents) will defy precise determination due to electro-chemicalsynergy. Hence, the electro-chemical measurements of complex organicsolutions, such as blood, necessitate interpretation of the signal dueto the lack of precision in identifying the contribution of a particulartargeted substance. Where precise measurements are required, theambiguity stemming from such interpretation is, at best, risky and, atworst, lethal. Moreover, the contribution of drift by both the referenceelectrode and the specimen electrode coupled with the junction potentialidentification problem, could lead to anomalous measurements.

Moving now to a practical problem associated with prior art systems, itis the manufacture and supply of both species specific electrodes andreference electrodes. Generally, the reference electrodes are of a moresophisticated construction in order that they be reusable. Without more,it is evident that measurements using such electrodes would differ inevery instance due to manufacturing tolerances. Accordingly, not onlyare the drift and calibration problems extant, but, also,standardization is difficult especially when conducting the severalmeasurements of different solutions required for calibration and unknownsolution evaluation.

Most analytical systems are exposed to the ambient environment. They arenot anaerobic. An anaerobic environment is desirable, first, to moreclosely match in vivo conditions. Furthermore, it is important, forexample, in blood gas analysis to avoid sample contamination from air soas to avoid skewing the results. Lastly, to obtain a series of substancemeasurements from a sample, requires considerable time and manyindividual measurements. Not only is the time factor detrimental but,also, specimen contamination and chemical changes in the specimen arelikely to occur. Hence, it is desirable to maintain an anaerobicmeasuring environment to achieve accurate measurements of certainsubstances, and most notably, blood gas concentrations. Lastly, mostknown systems do not contemplate fixing or providing fixed volumedelivery. Elaborate stirring or mixing arrangements are used to insureuniform transport to the sensor. It would be desirable to conductmeasurements of a fixed volume of solution and especially desirable toprovide analysis requiring only a small volume of solution uniformlydelivered to the sensor to make the measurements.

Other practical considerations arise relative to laboratory use by theclinician. In the event that a system is intended to be reusable, it isincumbent upon the operator or technician to insure that the electrodesare not contaminated when preparing for a test. Thorough cleaning andrecalibration is necessary for each use. Such efforts requireconsiderable labor and render cost ineffective the use of reusablesystems especially in hospital laboratories, etc. Where disposablesystems are employed, problems arise relating to the technician'stechniques.

Another aspect of electro-chemical apparatus that has escapeddevelopment is a compact, simply employable, field or laboratory useinstrument which can be operated by persons having a minimum of skilledtraining. Miniaturized and standardized equipments are not available forproviding analytical electro-chemical measurements like those describedabove.

SUMMARY OF THE INVENTION

It is, therefore, an object of this invention to overcome the problemsexperienced with the use of prior art techniques and methods.

It is another object of this invention to provide a method forgenerating accurate and reproducible substance concentrationmeasurements.

Still another object of this invention is to provide a method andapparatus for electro-chemical measurement requiring single pointcalibration for potentiometric, potentiostatic or resistivity analysis.

Still another object of this invention is to provide method andapparatus for electro-chemical substance concentration determinationswith a minimum cost and a minimum of effort by eliminating therequirement for a reference electrode.

It is another object of this invention to measure true activities oftargeted species in biological solutions without interference fromjunction potential errors caused by a reference electrode.

Yet another object of this invention is to provide techniques andapparatus for speedy measurements to avoid time-dependent internalchanges, to maximize stability and to minimize potential contaminationof the sensors.

It is another object of this invention to provide techniques andapparatus minimizing potential technician error and avoiding the needfor technician interpretation.

Still another object of this invention is to provide apparatus forsolution analysis which maintains the solution in an anaerobicenvironment.

Another object of this invention is to provide delivery of a fixedvolume of solutions for measurement.

Still another object of this invention is to provide measuring methodsand apparatus equally applicable to a range of analytical purposes suchas electro-chemical and optical measurements.

Yet another object of this invention is to provide techniques andapparatus capable of miniaturization and which is capable of employingan array of sensors for real-time, multispecies solution analysis.

Another object of this invention is to provide a cartridge which isdisposable or capable of reuse.

A further object of this invention is to provide a modular cartridgesystem where different cartridges for different measurements may besequentially introduced to signal processing apparatus.

It is another object of this invention to provide a universal sensorcartridge capable of incorporating a large number of different sensorsfor a broad range of different analytical techniques.

These and other objects are satisfied in part by a method for singlepoint calibration measurement of at least a first and a second speciesin solution employing at least a first, second and third sensors wherethe first sensor is sensitive to the first and second species, thesecond sensor is sensitive to the first species and the third sensor issensitive to the second species. The method contemplates contacting thesensors with a solution containing the first and second species,obtaining first and second signals where the first signal is thedifference between said first and second sensors and said second signalis the difference between said first and third sensors. The signals arethen conveyed to a signal processor. The next step involves contactingthe sensors with a second solution containing known quantities of thefirst and second species and obtaining third and fourth signals from thefirst and second sensors and said first and third sensors, respectively,which are conveyed to a signal processor, establishing algebraicconstants from said third and fourth signals, inputting the constantsinto a calculating device determining the concentration of said firstand second species. Equivalently, the technique can be performed byfirst introducing the known solution and the unknown solutionsubsequently. Also, as should be apparent to the skilled artisan, theconcentration determination is the equivalent of determining theactivity of the particular substances in solution. Summarizing thebenefits provided by this technique, in electro-chemical procedures, iteliminates the need for a reference electrode and corresponding slopeand intercept variations. It is readily adaptable to miniaturization. Itrequires only comparative measurements between determined species andonly N+1 sensors for measurement of N species. Moreover, it minimizeslabor and interpretation errors, especially when combined withequipments described herein.

Still other objects of this invention are satisfied by a cartridge forfacilitating analytical measurement of a solution, comprising a housingand a chamber for containing a predefined volume of solution, thechamber having a first end and a second end and being disposed withinthe housing. Combined with the chamber are an inlet port in fluidcommunication therewith which is located proximate to said first chamberend, and a waste reservoir of preselected volume in fluid communicationwith the chamber. Within the chamber is a means for minimizing fluidback-flow from said reservoir to said chamber and a sensor elementdisposed in the housing and interfacing with the chamber at apreselected location between the first end and the back-flow minimizingmeans. Lastly, the cartridge has a means for conveying signals generatedby the sensor through and out of the housing.

This cartridge is preferably constructed for a miniaturized instrument,maintains the test solution in an anaerobic environment, requiresintroduction of only a small amount of solution for test procedures, isadapted for incorporation of a number of different sensors and sensortypes and even contemplates disposability.

Still further objects of this invention are satisfied by providing asensor for evaluating a species in solution. The sensor embodies aconductive element capable of conducting signals having a first surfaceof particular cross-sectional dimensions coupled with species specificreactive means for reacting with a selected active species in solution.The reactive means is in intimate contact with the conductive elementand capable of generating a signal corresponding to the active speciesin solution. The reactive means is sized to cover the first surface andextend a substantial distance beyond the perimeter of the first surfaceto minimize edge effects.

The sensor arrangement, stated positively, facilitates uniform andreproducible measurements of a solution by insuring uniform interactionbetween the species specific receptor and the signal conductor at theinterface of the receptor and conductor. This is accomplished byeliminating or minimizing edge effects at the conductor perimeter. Thesensor is contemplated for incorporation in the equipments describedherein and is readily adapted for use with the referenceless technique.The critical teaching of the sensor structure is contrary to popularbelief. That is, it is not a precise sensor geometry which providesuniform measurement particularly for single point calibration proceduresbut the provision of a substantial overlap that minimizes edge effectcontributions and enhances measurements of greater precisionirrespective of the sensor geometry.

Certain of the objects stated above are finally satisfied by a compactinstrument for solution analysis. The instrument includes a housing, aninformation display means contained on a surface of the housing fordisplaying information, selection means for selecting the information tobe displayed, a receptacle of predetermined dimensions positioned on thehousing, and means for processing electrical signals and conveying theprocessed signals to the information display. The instrument furtherincludes a sensor containing cartridge for sensing properties of asolution and generating signals corresponding to the sensed properties.The cartridge is dimensioned to fit in said receptacle.

The described instrument is of a nature to provide a far simpler,miniaturized, easily handled analytical tool especially suited for fielduse. It contemplates minimizing the degree of clinical skill, knowledgeand labor required to operate and obtain accurate solution analysis.When combined with the methods and apparatus described herein, theinstrument is especially suited to provide a wide variety of analyticaldeterminations employing a host of different sensors and sensor types toachieve rapid sample evaluations.

In summary, the present invention provides a new referencelessanalytical method, a new sensor structure eliminating contribution fromedge effects, a cartridge which, among other aspects, is adapted forminiaturization and maintaining the test solution in a neutralenvironment, and a compact, self-contained, readily-employed analyticalmeasurement processing unit.

Given the foregoing summary, the skilled artisan will more readilyappreciate the advance this invention provides from review of thefollowing detailed description of the illustrated embodiments.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is a schematic representation of a sensor arrangement of theinvention.

FIG. 2 is a graphical depiction of flux density distribution acrosssensors.

FIG. 3 is a perspective view of a sensor cartridge of the invention.

FIG. 4 is a top view of a sensor array.

FIG. 5 is a perspective view of a compact instrument of the invention.

FIG. 6 is a view of the instrument in a folded configuration.

FIG. 7 is a perspective view of a disposable, sensor-containingcartridge.

FIG. 8 is a graphical representation of the sensor construction and fluxdensity variations caused by edge effects.

DETAILED DESCRIPTION OF THE ILLUSTRATED EMBODIMENTS

For organizational purposes, the illustrated embodiments will bedescribed first by the new method in the context of electro-chemicalanalysis of specified substrates; secondly, by the sensor structure inthe form of an electrode; thirdly, by a sensor assembly in the form of aminiaturized electrode containing cartridge; and lastly, a miniaturizedmicroprocessor based and solar powered instrument for field orlaboratory use.

At the outset, it should be noted that the illustrated embodimentscontemplate precise structures and miniaturization which are notnecessary for practice of certain aspects of the invention. For example,it is evident that laboratory equipment of considerable size can beconstructed. Also, multiple purpose cartridges incorporating referenceelectrodes can be provided each which embodies certain conceptsdescribed herein. Accordingly, it is not intended that the invention beso limited to the specific recitation below.

THE METHOD

Turning now to the method and referring to FIG. 1, it depictsmulti-channel sensor system 10. Sensor 10 features an array of sensors12 composed in this case of four individual electrode sensors, 16, 18,20 and 22. For the purpose of illustrative simplicity, electroactivesensor 16 is deemed to be sensitive to a species A, sensor 18 issensitive to a species B, sensor 20 is sensitive to species A and B, andsensor 22 is sensitive to species A and C. (These species may be chosenfrom many species such as potassium, sodium, chlorine, hydrogen ion, orselected biological and organic molecules.) All of species A, B and Care substances contained in a fluid which is to be electro-chemicallyevaluated using one of the two variations of the below-described method.More particularly, it is anticipated that a complex biological fluidsuch as blood will be subject to such an evaluation.

Although described in greater detail below, species specific coveringmembranes 17, 19, 21, and 23, corresponding respectively to sensors 16,18, 20 and 22, are impregnated with ion selected materials where theelectrodes are sensitive to A, B and C and combinations A and B as wellas A and C, respectively. The membranes and sensors are so arranged toprovide for a substantially uniform electrical signal caused byinteraction between the target species in solution and the electroactivecompound in the membrane. A corresponding charge develops between themembrane and the sensor thereby generating a charge distribution andpotential proportional to the ionic activity of the species.

The principal variant of the inventive technique is now described withreference to potentiometric electrode sensors 16, 18 and 20. It shouldbe appreciated by the skilled artisan that sensors 16, 18 and 20represent half-cells where the combination of two half-cells provide anelectro-motive force (EMF), representative of the potential differencebetween each of the respective sensors. Turning first to sensor 20, itis a combination electrode for species A and B, its electricalpotential, in simplest form, is expressed by the equation

    E.sub.20half =M.sub.A logC.sub.A +M.sub.B logC.sub.B +I.sub.AB (2)

M_(A) and M_(B) are constants for species A and B, respectively, whichfor particular compositions and electrodes, can be predetermined andprogrammed into a calculation device. C_(A) and C_(B) are theconcentrations of species A and species B, respectively. Equation 2 canbe further reduced to the expression:

    E.sub.20half =M.sub.AB [logC.sub.A +logC.sub.B ]+I.sub.AB  (3)

where the quantity of the electroactive species impregnate into membrane21 is carefully proportioned. For each such combination, beforemanufacture, it would first be necessary to evaluate the amounts toestablish the most effective combination to obtain the simpler equation.

Moving now to the other electrodes, the electrical potential of sensor18 which is sensitive to species B is expressible by the equation

    E.sub.18half =M.sub.B log[C.sub.B ]+I.sub.B                (4)

Likewise, the half-cell potential of sensor 16 specific to species A isexpressible as

    E.sub.16half =M.sub.A log[C.sub.A ]+I.sub.A                (5)

To one of ordinary skill in the art, the foregoing equations representthe classical Nernst-type equations obtained from ion selectiveelectrodes for measurement against standard electrodes. The instantinvention, however, eliminates the need for a reference electrode andits contribution to the signal. The elimination of the referenceelectrodes is accomplished by establishing cells between sensors 16 and20 and sensors 18 and 20, conveying the signals over wires 34 tomultiplexer 26 which is commanded over wires 36 by microcomputer 32. Thesignals are sent to op-amp 28; in this case a differential amplifier,where signals from 16 and 18 are passed to analog/digital converter 30and, ultimately, to microcomputer 32. The differential potentialscorresponding to E₂₀ -E₁₈ and E₂₀ -E₁₆ are thus obtained. Thedifferential signals are expressed by the equations ##EQU1##

Put more simply

    E.sub.20-18 =M.sub.A log[C.sub.A ]+I.sub.AB -I.sub.B       (7)

Correspondingly,

    E.sub.20-16 =M.sub.B log[C.sub.B ]+I.sub.AB -I.sub.A.      (8)

The slope values M_(A), M_(B), M_(AB) and any other slope constants areknown from prior testing of the particular electrode structures withstandard solutions. These values are either inputted or stored inmicrocomputer 32 for inclusion into the equations. Therefore, with theslope values and the signal values known, the constants and theconcentration values are determinable given measurement of a referencesolution to determine the constants. In order to solve the equations, areference solution having known concentrations of species A and B ismeasured. Since the constants I_(A), I_(B) and I_(AB) are the same forboth solutions, their contribution to the equation is subtracted out:##EQU2## Knowing the signal potential and the slope (M) values permitsdirect calculation of C_(A) and C_(B). By the foregoing, to obtaindifferential measurements of two distinct species requires only threeelectrodes; one electrode being selective for the combination of bothspecies being tested and, then, two individual electrodes selected toeach of the selected species being tested. Viewed simplistically, thecombination electrode provides a signal corresponding to the activity ofA +B where if the contribution of species A is substracted from thesignal, the concentration of B is determined. Correspondingly, where thecontribution of species B subtracted from the combinational electrodevalue, concentration of species A is determinable.

A second method for analysis of a greater number of species, can bepracticed by the invention. Employing the foregoing principles, theconcentration of a third species, C, may be determined by employing, atminimum, the fourth combination electrode 22 sensitive to species A andC. The concentration of C is determined by subtracting the signalproduced by electrode 16 from electrode 22. In such a case thecalibration solution must also include species C.

It should now be readily appreciated that the inventive method requiresonly one additional electrode to the number of species being evaluated.Mathematically, if N is the number of species targeted for analysis,only N+1 sensors are required to practice the technique. Moreover, thetechnique requires measurement of only two species containing solutions,the calibrant and the unknown solution.

A multiple combination system, as described above in the secondembodiment, may exhibit some interference due to the presence ofadditional species (B) in solution. Accordingly, it may proveadvantageous to have additional electrodes sensitive to species C aloneor/and the combination of A, B and C. In such a case, the calculationapparatus are employed to provide comparative data between the speciesspecific electrodes 16, 18 and 22 or the combination electrodes 20 andone sensitive to species A, B and C. Since multiple combinationelectrodes (more than two species) may be subject to electro-chemicallysynergistic interaction, anomalous signals may result. Hence, it issuggested that each electrode's sensitivity be limited to two species.

In summary, this invention permits evaluation of a solution for (N)separate species requiring only two measurements, the unknown solutionand the calibrant solution, using only (N+1) electrodes.

Some aspects of the above-described technique should now be underscored.Principally, in the context of electro-chemical analysis, the methoddispenses with the need for a reference electrode and, therefore,eliminates considerations for junction potential. Furthermore,elimination of the reference electrode minimizes "drift" problems byreducing the drift occurrence to two similarly structured electrodes.Rather than exhibiting relative combined drift of both the reference andspecies specific electrode each contributing its own district drift dueto dissimilar geometries, compositions, etc., employing similarlystructured and composed electrodes provides comparatively uniform drift.Hence, the drift component is often negligible or linear and assessable.It is not exponential and difficult to assess. (Drift is squared due tothe separate contribution of reference and species electrodes.)Secondly, the method lends itself to use in miniaturized devices.

It should be evident to the skilled artisan that not only does theinstant method provide a labor saving technique for multicomponentelectro-chemical analysis but also is an expedient for renderingreal-time results when needed. These benefits are especially importantin a clinical chemical environment during sensitive procedures such assurgery on a human patient.

ELECTRODE STRUCTURE

Conventional electrodes may be used with foregoing techniques and in thebelow-described apparatus. For example, wire, wire coated, and filmelectrodes, thick or thin film electrodes of a redox, semiconductor ortype involving a polymeric matrix immobilizing an electrochemicallyactive receptor impregnated therein, can be used. More specifically,variants of the thin film electrode described in U.S. Pat. No.4,214,968, the graphite electrode described in U.S. Pat. No. 4,431,508and the convex-domed electrode described in U.S. Pat. No. 4,549,951, mayall be employed in the arrangements and methods described herein and forthat reason are incorporated by reference.

The modification of the foregoing electrodes involves the selection ofthe ion selective electrode portion or membrane having a substantiallygreater cross-sectional area than the underlying conductor in order topromote uniform charge density between the solution and the conductor.

It has been suggested previously (see U.S. Pat. No. 4,549,951) that aconvex geometric configuration of the membrane contributed to promoteuniformity of signals from transport of the electroactive species of anion selective membrane to the interfacing cross-section of the conductorand, thus, accuracy and reproducibility of measurements. The dome-shapedmembrane electrode was conceived of for this purpose. However, what wentunrecognized is the contribution of edge effects to space chargedistribution and transport phenomena and, hence, (adherence from surfacetension, greater electron transfer, etc., generated along the perimeterof the conductive body) to the signal. Basically, edge effects resultfrom nonuniform layers of charge distribution between the interfaces ofsolution, the membrane and the electrically conductive member of theelectrode. The nonuniformity is particularly pronounced along theperimeter of the conductor and membrane due to surface phenomena andexposure to a relatively greater volume of solution with a correspondinghigher density of flux. This factor gives rise to slope variations fromelectrode to electrode even for the same species.

It has now been found that the elimination of edge effects promotessignal uniformity without a need to restrict the configuration of themembrane to a particular geometry. Accordingly, it is now believed to beno need for the membrane to possess any particular geometricconfiguration (dome shape, etc.) but rather to provide an area ofsufficiently greater size than the conductor cross-section to minimizeedge effects. Indeed, it is preferred to provide a membrane surface areahaving at least approximately twice the size of the cross-sectional areaof the conductor. However, more precisely, the degree of membraneoverlap is mathematically accessible from the membrane/electrodegeometry and classical electron transport equations.

Referring briefly to FIG. 2, it graphically depicts the electronpathways between solution S, rectangularly cross-section membrane 37,and domed membrane 38 to underlying conductors 36. Although some signalcontribution occurs from the outer membrane portions, the predominateuniform flux distribution is generated from the portion overlying theelectrode and a bevelled portion of between 30° to 45° flaring from theedge of conductor 36. To promote uniformity of edge effect and,therefore, avoid nonuniform measurement, the membrane area is increasedto extend well beyond the perimeter of the conductor.

Turning briefly to FIG. 8, it represents the contribution of edgeeffects to various electrode structures. Electrodes 110, 112 and 114,having concave, convex and flat membranes, respectively, eachdemonstrate uniform flux density across the entire conductor surface.Convex domed electrode 116, having a membrane extending a little beyondthe conductor perimeter, exhibits a small deviation in flux density.Electrode 118, with no extension exhibits considerable flux densityvariation across the conductor surface. As stated above, FIG. 7underscores the fact that this invention contemplates theminiaturization of nonuniform flux density along the edge of theelectrode by providing a membrane of considerably greater cross-sectionthan the underlying conductor surface. Hence, the instant inventioncontemplates that the electrode will contain a species reactive portionor membrane having an overlap so as to possess a solution interface areaconsiderably greater (approximately twice) than the cross-section of theconductor.

In summary, the electrodes contemplated for use in the instant inventionare known electrodes modified to provide an increased electro-chemicallyactive surface of considerably greater surface contact area than theunderlying electrically conductive portion of the electrode to eliminateedge effects and corresponding uneven flux density.

THE CARTRIDGE

The contemplated sensor containing cartridge is intended to containseveral microsensors of similar or different types, maintain ananaerobic sample chamber, and provide a fixed value delivery means tothe fixed volume chamber.

Referring now to FIG. 3, it depicts cartridge 40 comprising twoprincipal sections, chamber housing 42 and lower insert portion 44.Contained within chamber housing 42 is fixed volume chamber 46 designed,typically, to hold a volume of less than one milliliter and preferablybetween 10-50 microliters. Chamber 46 is generally of a rectangularconfiguration and is sealed within chamber housing 42. Within chamberhousing 42, sensors 16, 18, 20 and 22 are embedded and disposed in anarray on the lower surface of chamber 46. The sensors are electricallyisolated from each other and are positioned in chamber 46 in a mannerwhere fluid introduced therein will completely cover membranes 17, 19,etc.

Disposed transversely along one side of chamber 46, is vented wastereservoir 50 having a volume capacity of 4-6 times that of chamber 46.One direction flow vents 52 are provided at selected locations in orderfor air or gas to escape and allow fluid from chamber 42 to evenly flowinto and fill reservoir 50. Between reservoir 50 and the sensors arelocated furrow 48 and weir 43. Furrow 48 and weir 43 are designed toprevent fluid back-flow from waste reservoir 50 into chamber 46 havingdisposed on its lower surface the array of sensors. Especially whereintended for use in the field, weir 43 should be of a height exceedingthe thickness of membranes 17, 19, etc. to maintain the test solutionthereover. Furrow 48 and weir 43 serve to prevent fluid back- flow fromwaste reservoir 50 into chamber 46 and resulting mass transfer andcontamination between the waste fluids and the analytical fluid. It isnoted that the weir may be unnecessary where the cartridge is of adesign to take advantage of surface tension to stabilize the samplefluid, on the one hand, and the calibrant fluid, on the other hand, overthe sensor.

At the opposite end of chamber housing 42 from waste reservoir 50 arecalibration fluid input port 56, calibration fluid syringe 54 andspecimen inlet port 62 with specimen input element 60 extendingtherefrom. Specimen input element 60 provides a rubber septum across itsupper surface for injection of the specimen into element 60 through port62 and into chamber 46 from a conventional syringe or, alternatively, acapillary tube. Although it is desirable to inject an amount of specimenfluid equal to the volume of chamber 46, any excess will flow intofurrow 48 and, subsequently, into waste reservoir 50.

Calibration fluid syringe 54 contains a predetermined volume of anappropriate calibration fluid containing substances for which thesensors arrayed within chamber 46 are sensitive. Preferably, acontrolled volume of calibration fluid is injected into chamber 46 bydepressing plunger 58 where the fluid flows through input port 56 andinto the chamber.

Moving now to the structure lower insert portion 44, like chamberhousing 42, it is preferably composed of a suitably rigid, strong,transparent polymer having the conductive elements (graphite, wire, etc)from sensors 16, 18, etc., extending through its entire length.

By providing significant sensor elongation, especially whenelectro-chemical measurements are performed, the elongation minimizessignal interferences from adjacent sensors. As a practical matter,during manufacture of a membrane covered electrode, the membrane isdeposited over the conductor in a partially gelled condition. Theremaining solvent, generally organic, is then evaporated. However, somesolvent will migrate into pores in the cartridge body. Migrating solventcan carry with it the electroactive species. Hence, the cartridge body,itself, may be sensitized or even cross-sensitized. Where electrodes arepositioned very close to each other, cross-contamination can occur.Thus, a species specific electrode may generate a small signalcorresponding to another species for which the neighboring electrode issensitive. This possibility is enhanced when the cartridge body is veryshort, the degree of migration is correspondingly reduced, andintermingling occurs close to the sensor receptor surface. By elongatingthe sensors and cartridge, gravity causes the solution bearing, residualelectroactive species to follow a downward path adjacent the electrodeinstead of transverse intermingling a short distance from the receptormembrane. Hence, it is preferred that the cartridge be of sufficientlength to minimize such effects.

Returning to the structure of cartridge 40, waste reservoir 50 extendstoward the bottom of lower insert portion 44. Projecting from the bottomof chamber 44 are electrical point contacts 47 which provide electricalcommunication between sensors 16, 18, and an appropriate signaldetector. Due to potential internal signal interference or interferencefrom external electrical noise, it may be desirable to insulate each ofsensors 16, 18, etc. Accordingly, sensor 16 is illustrated withinsulative sheathing 49 disposed therearound. If all the sensors are soinsulated, the opportunity for electrical signal interference isminimized.

In brief, cartridge 40 is used by injecting a sufficient volume of thespecimen fluid into chamber 46 via inlet port 62 to fill chamber 46.Measurements of the electro-chemical activity are made via the array ofsensors. Once measurements are taken, a fixed volume of calibrationfluid is introduced via syringe 54 through input port 56 which washesthe specimen fluid from chamber 46 into furrow 48 over weir 43 and intowaste reservoir 50. A second portion is added which washes the firstportion out of chamber 46 over weir 43 and into reservoir 50. Finally, athird portion is added to displace the second portion. By this means,residual specimen fluid is substantially completely removed from chamber46 and electro-chemically active membranes 17, 19, etc. Furthermore, byproviding multiple washings, if the specimen contains a higherconcentration of a particular ion than the calibration fluid, themultiple washings permit an equilibrium to be established to minimizeinaccurate measurements of a particular ion concentration in thecalibrant solution due to residual ionic activity from the specimen onmembranes 17, 19, etc.

In FIG. 4 is illustrated an alternative embodiment of cartridge chamberhousing 42 and chamber 46. In this embodiment there is an array offourteen sensors which have the capacity for analysis of as many asthirteen electro-chemical active species. At one end of chamber 46, likethe embodiment in FIG. 3, is disposed waste reservoir 50 for containingthe analyzed specimen sample and the volume of calibration fluidemployed for washing out the specimen fluid from chamber 46. Betweenreservoir 50 and the array of sensors is disposed furrow 48 and weir 43.Weir 43 in this case is positioned between the furrow and the sensorsand assists to define a specific volume of fluid that will be containedwithin chamber 46. The fluid introduction may follow the steps describedabove or, alternatively, the calibrant may first be introduced intochamber 46 and measurements taken followed by introduction of thespecimen solution into the chamber with measurements being taken of thespecimen fluid. Where the calibrant is first introduced, it is possibleto eliminate particular washing steps by providing a relativelysubstantial volume of specimen fluid to displace the calibrant solution,develop an equilibrium and be subject to measurement. Any excessspecimen fluid will flow into waste reservoir 50.

The above-described cartridge embodiments are contemplated as beingdisposable as they would be composed of a relatively, inexpensivepolymeric material. However, it is also possible to reuse the cartridge,given the inclusion of reusable sensors within the cartridge, byproperly cleaning and otherwise freeing cartridge 40 from contamination.As would be expected in such an embodiment, reservoir 50 would beprovided with an appropriate fluid outlet near or at the bottom of thereservoir in lower insert portion 44 in order to permit a series ofappropriate washings. Another alternative construction would be toprovide an open-topped fluid containing chamber 46. This, in certaincases, would be undesirable as it would eliminate the anaerobicenvironment by exposing the species and calibrant to an ambientatmosphere. (As noted above, particularly in the context of biomedicalmeasurements, it is preferred to maintain an anaerobic environment.) Forthis reason, it is suggested that chamber 46 and waste reservoir 50 beflushed with a neutral gas such as nitrogen following construction andprior to use to minimize the presence of atmospheric oxygen and carbondioxide during testing.

An additional construction variant includes modifying element 60 to be aflow diversion valve or dispenser adapted to extract a sample directlyfrom the source. For example, element 60 may be combined with a catheterto extract blood directly from a patient's body. Point contacts 47 mayalso be modified both in structure and position. They may exit lowerportion 44 on its side and be of a structure to establish wipingelectrical contact with an appropriate mating receptacle.

Lastly, it is possible to modify the cartridge for instruments otherthan electro-chemical analyzers. For example, optical fibers could beincorporated for measurement of fluid optical properties. In this case,it would be suggested to have source fibers and receptor fibers disposedin an array to maximize optical transmission and reception. Preferably,conventional available coaxial fibers would be used. Moreover, the uppersurface of chamber 46 can be coated with an optically reflectivematerial. An additional variant would be optical colorometric analysisof the covering membrane impregnated with a species specific interactivesubstance which undergoes a color change upon reaction. Color changescan be detected using coaxial optical sensors. As one further variant,optical and electrochemical sensors can be combined in the samecartridge.

In summary, cartridge 40 serves the function of positionally stabilizingand maintaining a specific geometry between the sensors housed therein,defines a precise volume of fluid for analysis, provides an anaerobictesting environment, avoids sensor contamination, provides wastecontaminant while avoiding fluid intermingling and means for precisealignment of the sensors with appropriate detection apparatus. Moreover,it is adaptable for use with a host of conventional sensors, forexample, potentiometric, potentiostatic, resistance, colorometric, etc.,analysis.

MINIATURIZED SYSTEM

In FIGS. 5 and 6 are depicted a miniaturized instrument for use of asensor containing cartridge, like that described above, containingelectrodes, like those disclosed above, and contemplating measurementsby the analytical techniques set forth above. Compact instrument 80dedicated for biomedical use, is comprised of fold-up case 82 featuringupper portion 81 and fold out portion 83 which are hinged (not shown).The electronic components employed within the case are commerciallyavailable. They are microprocessors, random access memories (RAM)s, readonly memories (ROM)s, amplifiers, switches, analog to digitalconverters, power capacitors, transformers, etc.

The principal features of upper portion 81 are liquid crystal displaypanel 84, controlled by the microprocessor (not shown) and a row ofactuation buttons 86 for activating the particular function desired.Upper portion 81 is also provided with an appropriately sized cartridgereceptacle (not illustrated) to permit insertion of sensor cartridge 94therein. Once inserted, as described above, the electrodes contained bycartridge 94 are processed and are displayable on the liquid crystaldisplay screen. Buttons 86 facilitate selection of the desired, W forexample, particular blood gas concentrations or even blood pressure, ondisplay 84. The receptacle can be modified to include an opticalcharacter recognition device or magnetic pickup device for readinginformation placed on the side of cartridge 94. For example, a bar codeor piece of magnetically encoded tape can be positioned on the cartridgewhich would automatically input data such as slope values (see methodabove), identify specific sensors and combination sensors, etc. Themodification would eliminate the need for the operator to input suchvalues and information by, for example, a programming keyboard (notillustrated). Furthermore, the codes could reprogram buttons 86 forspecific tests performed by specific cartridges.

Lower panel 83 features a solar power panel 89, RS232 port 90 andplug-in adaptors 91 and 92 for connecting peripheral equipment such as aphonocardiogram and blood pressure monitor. Signals from aphonocardiogram or blood pressure monitor are displayable on the LCDfollowing appropriate signal processing by the microprocessor andactivation by the appropriate button. RS232 port 90 permits digitalcommunication between the unit and a remote digitalized patientinformation storage area should it be desirable to convey the dataprocessed from cartridge 94 or from ancillary equipment such as theabove-stated phonocardiogram or blood pressure monitor to a computer,etc. Due to the advances in microprocessor and electronics technology,in addition to the miniaturization provided by the structures andtechniques defined above, it is possible to provide a physicalembodiment of instrument 80 adapted to fit into a pocket. As such thedimensions should not be in excess of 3 1/4 inches wide, 9 inches long,and 1 1/4 inches deep. Furthermore, the weight of the entire unit can berestricted to approximately one-half pound. Hence, the unit is easilyhandled and stored. Indeed, it is possible for a doctor to slip theentire unit, when folded, as depicted in FIG. 6, into his coat pocketfollowing patient examination. Moreover, given the provision of solarpanel 89 for generation of needed power, it is not necessary for thephysician, medical technician or clinician to have an electrical poweroutlet readily available. Alternatively, chemical batteries, etc., canbe incorporated as an appropriate power source. Thus, the unit isreadily adapted to use in the field, as for example, at accident scenes,etc. The RAMs incorporated in instrument 80 permit the medicaltechnician or doctor to make a series of patient samplings which can belater recalled and inputted into a primary patient data bank.

Now turning to FIG. 7, a portable, disposable variant of cartridge 40,described above and contemplated for use with unit 80, is illustrated.Primarily, cartridge 94 includes upper portion 95 and lower portion 97where lower portion 97 is adapted to be inserted into the complementaryaperture provided in unit 80 and establish electrical contacttherebetween. It is contemplated that appropriate electronic circuityand control like that described in reference to FIG. 1 is incorporatedinto instrument 80 to provide fluid analysis by the method described.The ROMs employed in such a unit, for example, would hold slopeinformation for particular substances relative to the particularelectrode structure.

Moving now to the particular configuration of upper portion 95, itincludes a chamber containing calibrant solution and push-buttoncalibrant injector 98 for flooding the specimen chamber (notillustrated). Further illustrated is specimen port 96 for injection ofblood or other appropriate fluid into the specimen chamber. In thisembodiment, it is contemplated that the waste reservoir be entirelycontained within upper portion 95. The operation of this cartridge isidentical to the procedures described earlier in this application.

As is readily apparent, pocket-sized unit 80 and cartridge 94 aredirected to use by medical personnel, either in a hospital environmentor in the field. Of course, the same principles may be employed inalternative disciplines such as environmental aquatic analysis.

Given the foregoing description of the system, the cartridge, themodified electrode structure and the single calibration referencelesstechnique, a host of modifications and variations thereto should now beapparent to one of ordinary skill in the art. It is intended that suchmodifications and variations fall within the scope of the invention asdescribed by the appended claims.

I claim:
 1. A method that is capable of single point calibrationmeasurement of at least first and second dissimilar species in solutionemploying only ion specific sensors, the ion specific sensors beingequal to the number of species plus one where in a multi-componentsolution containing at least two species to be measured a first sensoris sensitive to the first and second species, a second sensor issensitive to the first species and a third sensor is sensitive to thesecond species, the method comprising the steps of:(a) contacting thesensors with a solution containing the first and second species, (b)obtaining first and second signals, said first signal being thedifference between said first and second sensors and said second signalbeing the difference between said first and third sensors, (c) conveyingthe first and second signals to a signal processor, (d) contacting thesensors with a second solution containing known quantities of the firstand second species and obtaining third and fourth signals from the firstand second sensors and said first and third sensors, respectively, (e)conveying the third and fourth signals to a signal processor, (f)establishing algebraic constants from said third and fourth signals, (g)inputting the constants into a calculating device to determine theconcentration of said first and second species.
 2. A method according toclaim 1 where the signals represent electrode voltages.
 3. A methodaccording to claim 2 where the concentration of the first species isdetermined by calculations using the half cell voltage produced by thesecond sensor and the half cell voltage produced by the third sensor. 4.A method according to claim 3 where the voltages from the sensors aremultiplexed and passed to a differential amplifier.
 5. A methodaccording to claim 1 including the steps of providing the sensors with asubstantially uniform geometry at the solution interface and minimizingflux density.
 6. A method according to claim 1, further comprising thestep of minimizing flux density variations caused by edge effects.
 7. Amethod according to claim 1, further including the steps of arraying thesensors along a surface of a chamber isolated from the ambientenvironment and introducing a precise volume of solution into thechamber.
 8. A method according to claim 1 where the sensors are arrayedin a cartridge and the cartridge is electrically connectable to a signalprocessor further including the step of establishing electrical contactbetween the cartridge and the processor.
 9. A method according to claim1 where the number of species to be measured is N and the number ofsensors is N+1 where N is greater than
 2. 10. The method of determiningthe quantities of materials in a solution comprising the steps of thedeveloping a plurality of ion selective electrode potentials in a systemhaving only ion specific electrodes, said potentials being a function ateach electrode of the material of the electrode and of material in asolution in which the electrodes are immersed, anddeveloping signalssuch that the deference in the signals represent the differences in theEMFs of the reactions taking place only at the ion specific electrodes.